The long-term goal of this project is to understand the molecular mechanisms governing retinal development in vertebrates using zebrafish as a model. This application focuses on the development of retinal ganglion cells, which are important for normal vision and defects in which are involved in many human eye diseases including glaucoma. Recent results have demonstrated direct or indirect evidence for important roles of hedgehog (Hh) signaling, extracellular signal-regulated kinase (ERK) activity, and atonal homologue 5 (ath5), in retinal ganglion cell development. These pathways are known to be essential for development of the fly eye. Remarkably, the differentiation of retinal ganglion cells proceeds in a wave-like manner, reminiscent of progression of the differentiation of R8 photoreceptors in the fly eye. This application aims to determine the roles of and interactions among Hh, ERK, and Ath5 pathways in vertebrate retinogenesis. We will use zebrafish mutants that lack Hh signaling due to the disruption of smoothened (smo) gene, a component of Hh receptor, and a retrovirus-mediated gene expression system to manipulate the activities of these pathways in the retina. Information gained from this study will have important implications in understanding the evolution of the vertebrate retina and the mechanisms governing retinogenesis in vertebrates, including humans. The specific aims of this study are: 1) to determine whether Hh signal transduction regulates retinal differentiation in a cell-autonomous or nonautonomous manner in mosaics generated by reciprocal blastomere transplantation and by retrovirus-mediated retinal expression of either activated Smo or a specific inhibitor of protein kinase A (PKI) in smo mutant retina; 2) to determine whether the ERK pathway is essential for retinal ganglion cell differentiation and to identify the upstream regulators of ERK activity by expressing positive and negative regulators, and constitutively active and dominant-negative components of the ERK pathway; 3) to determine whether Hh signaling regulates ERK activity, and ERK in turn regulates ath5 activity in the developing vertebrate retina, and whether ath5-induced retinal ganglion cell differentiation requires Hh and/or ERK signaling.